2-substituted-2-imidazolines

ABSTRACT

The present invention relates to compositions and methods for inhibiting nonenzymatic cross-linking (protein aging) which contain novel 2-substituted-2-imidazolines. Accordingly, a composition is disclosed which comprises an agent capable of inhibiting the formation of advanced glycosylation endproducts of target proteins by reacting with the carbonyl moiety of the early glycosylation product of such target proteins formed by their initial glycosylation. The method comprises contacting the target protein with the composition. Both industrial and therapeutic applications for the invention are envisioned, as food spoilage and animal protein aging can be treated.

This invention was made in part with government support under Grant No.PHS AM 19655 awarded by The National Institute of Health. The governmenthas certain rights in the invention.

The present application is a continuation-in-part of copendingapplication Ser. No. 07/605,654, filed Oct. 30, 1990, now U.S. Pat. No.5,140,048, which is a division of U.S. Ser. No. 07/264,930 filed Nov. 2,1988, and now U.S. Pat. No. 4,983,604, which is a Division of U.S. Ser.No. 798,032, filed Nov. 14, 1985 and now U.S. Pat. No. 4,758,583, whichis a Continuation-In-Part of U.S. Ser. No. 590,820, filed Mar. 19, 1984and now U.S. Pat. No. 4,665,192. Applicants claim the benefits of theseApplications under U.S.C. §120.

RELATED PUBLICATIONS

The Applicants are co-authors of the following articles directed to thesubject matter of the present invention: "COVALENT ATTACHMENT OF SOLUBLEPROTEINS BY NONENZYMATICALLY GLYCOSYLATED COLLAGEN: ROLE IN THE IN SITUFORMATION OF IMMUNE COMPLEXES", Brownlee et al., J. Exp. Med., 158, pp.1730-1744 (1983); and "AGING OF PROTEINS: ISOLATION AND IDENTIFICATIONOF FLUORESCENT CHROMOPHORE FROM THE REACTION OF POLYPEPTIDES WITHGLUCOSE", Pongor et al., Proc. Natl. Acad. Sci. USA, 81. pp. 2684-2688,(1984), and "ADVANCED GLYCOSYLATION ENDPRODUCTS IN TISSUE AND THEBIOCHEMICAL BASIS OF DIABETIC COMPLICATIONS", Brownlee et al., The NewEng. J. of Med., 318, pp. 1315-1321 (1988). All of the abovepublications are incorporated herein by reference.

BACKGROUND OF THE INVENTION

The present invention relates generally to the aging of proteinsresulting from their reaction with glucose and other reducing sugars,and to the inhibition of the reaction of nonenzymatically glycosylatedproteins and the often resultant formation of advanced glycosylationendproducts and cross-links.

The reaction between glucose and proteins has been known for some time.Its earliest manifestation was in the appearance of brown pigmentsduring the cooking of food, which was identified by Maillard in 1912,who observed that glucose or other reducing sugars react with aminoacids to form adducts that undergo a series of dehydrations andrearrangements to form stable brown pigments Maillard, C.R. Acad. Sci.,154, pp. 66-68, (1912). Further studies have suggested that stored andheat treated foods undergo nonenzymatic browning as a result of thereaction between glucose and the polypeptide chain, and that theproteins are resultingly cross-linked and correspondingly exhibitdecreased bioavailability.

This reaction between reducing sugars and food proteins was found tohave its parallel in vivo. Thus, the nonenzymatic reaction betweenglucose and the free amino groups on proteins to form a stable,1-deoxyketosyl adduct, known as the Amadori product, has been shown tooccur with hemoglobin, wherein a rearrangement of the amino terminal ofthe beta-chain of hemoglobin by reaction with glucose, forms the adductknown as hemoglobin A_(lc). The reaction has also been found to occurwith a variety of other body proteins, such as lens crystallins,collagen and nerve proteins. See, Bunn et al., Biochem. Biophys. Res.Comm., 67, pp. 103-109 (1975); Koenig et al., J. Biol. Chem., 252, pp.2992-2997 (1977); Monnier et al., in Maillard Reaction in Food andNutrition, ed. Waller, G. A., American Chemical Society, 215, pp.431-448(1983); and Monnier and Cerami, Clinics in Endocrinology and Metabolism.11, pp. 431-452 (1982).

Moreover, brown pigments with spectral and fluorescent propertiessimilar to those of late-stage Maillard products have also been observedin vivo in association with several long-lived proteins, such as lensproteins and collagen from aged individuals. An age-related linearincrease in pigment was observed in human dura collagen between the agesof 20 to 90 years. See, Monnier et al., Science, 211, pp. 491-493(1981); Monnier et al., Biochem. Biophys. Acta, 760, pp. 97-103 (1983);and, Monnier et al., Proc. Natl. Acad. Sci., 81, pp. 583-587 (1984).Interestingly, the aging of collagen can be mimicked in vitro by thecross-linking induced by glucose; and the capture of other proteins andthe formation of adducts by collagen, also noted, is theorized to occurby a cross-linking reaction, and is believed to account for the observedaccumulation of albumin and antibodies in kidney basement membrane See,Brownlee et al, J. Exp. Med., 158, pp. 1739-1744 (1983); and Kohn etal., Diabetes 33 No. 1, pp. 57-59 (1984).

In Parent Application Serial No. 798,032, a method and associated agentswere disclosed that served to inhibit the formation of advancedglycosylation endproducts by reacting with the early glycosylationproduct that results from the original reaction between the targetprotein and glucose. Accordingly, inhibition was postulated to takeplace as the reaction between the inhibitor and the early glycosylationproduct appeared to interrupt the subsequent reaction of theglycosylated protein with additional protein material to form thecross-linked late-stage product. One of the agents identified as aninhibitor was aminoguanidine, and the results of further testing haveborne out its efficacy in this regard.

While the success that has been achieved with aminoguanidine and similarcompounds is promising, a need continues to exist to identify anddevelop additional inhibitors that broaden the availability and perhapsthe scope of this potential activity and its diagnostic and therapeuticutility.

SUMMARY OF THE INVENTION

In accordance with the present invention, a method and compositions aredisclosed for the inhibition of the advanced glycosylation of proteins(protein aging). In particular, the compositions comprise agents forinhibiting nonenzymatic cross-linking (protein aging) due to theformation of advanced glycosylation endproducts. The agents may beselected from those materials capable of reacting with the earlyglycosylation product from the reaction of glucose with proteins andpreventing further reactions. Cross-linking caused by other reactivesugars present in vivo or in foodstuffs, including ribose, galactose andfructose would also be prevented by the methods and compositions of thepresent invention.

The agents comprise compounds having the following structural formula:##STR1## wherein n is 1 or 2;

R₁ is an amino group or a hydroxyethyl group; and

R₂ is an amino, a hydroxyalkylamino, a lower alkyl group or a group ofthe formula

    alk-Y

wherein alk is a lower alkylene group and Y is selected from the groupconsisting of hydroxy, lower alkoxy, lower alkylthio, lower alkylaminoand heterocyclic groups containing 4-7 ring members and 1-3 heteroatoms;

with the proviso that when R is a hydroxyethyl group then R₂ is an aminogroup; their biocompatible and pharmaceutically acceptable acid additionsalts, and mixtures thereof; and a carrier therefor.

The compounds utilized in the compositions of this invention appear toreact with the early glycosylation product thereby preventing the samefrom later forming the advanced glycosylation end products which lead toprotein cross-links, and thereby, to protein aging.

The present invention also relates to a method for inhibiting proteinaging by contacting the initially glycosylated protein at the stage ofthe early glycosylation product with a quantity of one or more of theagents of the present invention, or a composition containing the same.In the instance where the present method has industrial application, oneor more of the agents may be applied to the proteins in question, eitherby introduction into a mixture of the same in the instance of a proteinextract, or by application or introduction into foodstuffs containingthe protein or proteins, all to prevent premature aging and spoilage ofthe particular foodstuffs.

The ability to inhibit the formation of advanced glycosylationendproducts carries with it significant implications in all applicationswhere protein aging is a serious detriment. Thus, in the area of foodtechnology, the retardation of food spoilage would confer an obviouseconomic and social benefit by making certain foods of marginalstability less perishable and therefore more available for consumers.Spoilage would be reduced as would the expense of inspection, removal,and replacement, and the extended availability of the foods could aid instabilizing their price in the marketplace. Similarly, in otherindustrial applications where the perishability of proteins is aproblem, the admixture of the agents of the present invention incompositions containing such proteins would facilitate the extendeduseful life of the same. Presently used food preservatives anddiscoloration preventatives such as sulfur dioxide, known to causetoxicity including allergy and asthma in animals, can be replaced withcompounds such as those described herein.

The present method has particular therapeutic application as theMaillard process acutely affects several of the significant proteinmasses in the body, among them collagen, elastin, lens proteins, and thekidney glomerular basement membranes. These proteins deteriorate bothwith age (hence the application of the term "protein aging") and as aconsequence of diabetes. Accordingly, the ability to either retard orsubstantially inhibit the formation of advanced glycosylationendproducts carries the promise of treatment for diabetes and, ofcourse, improving the quality and, perhaps, duration of animal life.

The present agents are also useful in the area of personal appearanceand hygiene, as they prevent the staining of teeth by cationicanti-microbial agents with anti-plaque properties, such aschlorhexidine.

Accordingly, it is a principal object of the present invention toprovide a method for inhibiting the extensive cross-linking of proteinsthat occurs as an ultimate consequence of the reaction of the proteinswith glucose and other reactive sugars, by correspondingly inhibitingthe formation of advanced glycosylation endproducts.

It is a further object of the present invention to provide a method asaforesaid which is characterized by a reaction with an initiallyglycosylated protein identified as an early glycosylation product.

It is a further object of the present invention to provide a method asaforesaid which prevents the rearrangement and cross-linking of the saidearly glycosylation products to form the said advanced glycosylationendproducts.

It is a yet further object of the present invention to provide agentscapable of participating in the reaction with the said earlyglycosylation products in the method as aforesaid.

It is a still further object of the present invention to providetherapeutic methods of treating the adverse consequences of proteinaging by resort to the aforesaid method and agents.

It is a still further object of the present invention to provide amethod of inhibiting the discoloration of teeth by resort to theaforesaid method and agents.

It is a still further object of the present invention to providecompositions including pharmaceutical compositions, all incorporatingthe agents of the present invention.

Other objects and advantages will become apparent to those skilled inthe art from a consideration of the ensuing description.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

In accordance with the present invention, agents, compositions includingpharmaceutical compositions containing said agents and associatedmethods have been developed which are believed to inhibit the formationof advanced glycosylation endproducts in a number of target proteinsexisting in both animals and plant material. In particular, theinvention relates to a composition which may contain one or more agentscomprising compounds having the structural formula ##STR2## wherein n isI or 2;

R₁ is an amino group or a hydroxyethyl group; and

R₂ is an amino, a hydroxyalkylamino, a lower alkyl group or a group ofthe formula

    alk-Y

wherein alk is a lower alkylene group and Y is selected from the groupconsisting of hydroxy, lower alkoxy, lower alkylthio, lower alkylaminoand heterocyclic groups containing 4-7 ring members and 1-3 heteroatoms;

with the proviso that when R₁ is a hydroxyethyl group then R₂ is anamino group; their biocompatible and pharmaceutically acceptable acidaddition salts, and mixtures thereof; and a carrier therefor.

The lower alkyl, lower alkylene and lower alkoxy groups referred toherein contain 1-6 carbon atoms and include methyl, methylene, methoxy,ethyl, ethylene, ethoxy, propyl, propylene, propoxy, butyl, butylene,butoxy, pentyl, pentylene, pentyloxy, hexyl, hexylene, hexyloxy and thecorresponding branched chain isomers thereof. The heterocyclic groupsreferred to herein include 4-7 membered rings having at least one and upto 3 heteroatoms therein. Representative heterocyclic groups are thosesuch as morpholino, piperidino, piperazino, methylpiperazino, andhexamethylenimino.

Equivalent to the compounds of Formula I for the purpose of thisinvention are the biocompatible and pharmaceutically acceptable saltsthereof. Such salts can be derived from a variety of organic andinorganic acids including but not limited to, methanesulfonic,hydrochloric, toluenesulfonic, sulfuric, maleic, acetic and phosphoricacids.

Of the compounds encompassed by Formula I, certain combinations ofsubstituents are preferred. For instance, when R₁ is a hydroxyethylgroup, then R₂ is an amino group. When R₁ is an amino group, then R₂ ispreferably a hydroxy lower alkylamino, a lower alkyl group or a group ofthe formula alk-Y, wherein alk is a lower alkylene group and Y isselected from the group consisting of hydroxy, lower alkoxy, loweralkylthio, lower alkylamino and heterocyclic groups containing 4-7 ringmembers and 1-3 heteroatoms.

Representative compounds of the present invention are:

1-amino-2-[2-(2-hydroxyethyl)hydrazino]-2-imidazoline;

1-amino-2-(2-hydroxyethylamino)-2-imidazoline;

1-(2-hydroxyethyl)-2-hydrazino-1,4,5,6-tetrahydropyrimidine;

1-(2-hydroxyethyl)2-hydrazino-2-imidazoline;

1-amino-2-([2-(4-morpholino)ethyl]amino)imidazoline;

1-amino-2-([3-(4-morpholino)propyl]amino)imidazoline;

1-amino-2-([3-(4-methylpiperazin-1-yl)propyl]-amino)imidazoline;

1-amino-2-([3-(dimethylamino)propyl]amino)imidazoline;

1-amino-2-[(3-ethoxypropyl)amino]imidazoline;

1-amino-2-([3-(1-imidazolyl)propyl]amino)imidazoline;

1-amino-2-(2-methoxyethylamino)-2-imidazoline;

1-amino-2-(3-isopropoxypropylamino)-2-imidazoline;

1-amino-2-(3-methylthiopropylamino)-2-imidazoline;

1-amino-2-[3-(1-piperidino)propylamino)imidazoline;

1-amino-2-[2,2-dimethyl-3-(dimethylamino)propylamino]-2-imidazoline;

1-amino-2-(neopentylamino)-2-imidazoline;

and the biocompatible and pharmaceutically acceptable salts thereof.

The above compounds are capable of inhibiting the formation of advancedglycosylation endproducts on target proteins. The cross-linking of theprotein to form the advanced glycosylation endproduct contributes to theentrapment of other proteins and results in the development in vivo ofconditions such as reduced elasticity and wrinkling of the skin, certainkidney diseases, atherosclerosis, osteoarthritis and the like.Similarly, plant material that undergoes nonenzymatic browningdeteriorates and, in the case of foodstuffs, become spoiled or toughenedand, consequently, inedible. Thus, the compounds employed in accordancewith this invention inhibit this late-stage Maillard effect andintervene in the deleterious changes described above.

The rationale of the present invention is to use agents which block thepost-glycosylation step, i.e., the formation of fluorescent chromophoressuch as that identified in Pongor, et al., supra, among others, thepresence of which chromophores is associated with, and leads to adversesequelae of diabetes and aging. An ideal agent would prevent theformation of the chromophore and its associate cross-links of proteinsto proteins and trapping of proteins on the other proteins, such asoccurs in arteries and in the kidney.

The chemical nature of the early glycosylation products with which thecompounds of the present invention are believed to react, isspeculative. Early glycosylation products with carbonyl moieties thatare involved in the formation of advanced glycosylation endproducts, andthat may be blocked by reaction with the compounds of the presentinvention, have been postulated. In one case, the reactive carbonylmoieties of Amadori products or their further condensation, dehydrationand/or rearrangement products, may condense to form advancedglycosylation endproducts Another proposed mechanism is the formation ofreactive carbonyl compounds, containing one or more carbonyl moieties(such as glycolaldehyde, glyceraldehyde or 3-deoxyglucosone) from thecleavage of Amadori or other early glycosylation endproducts (see, forexample, Gottschalk, A. (1972) in The Glycoproteins (Gottschalk, A., ed)Part A, pp. 141-157, Elsevier Publishing Co., New York; Reynolds, T. M.(1965) Adv. Food Res., 14, pp. 167-283), and by subsequent reactionswith an amine or Amadori product to form carbonyl containing advancedglycosylation products such as the alkylformylglycosylpyrroles.

Several investigators have studied the mechanism of advancedglycosylation product formation. In vitro studies by Eble et al.,(1983), "Nonenzymatic Glucosylation and Glucose-dependent Cross-linkingof Protein", J. Biol. Chem., 258:9406-9412, concerned the cross-linkingof glycosylated protein with nonglycosylated protein in the absence ofglucose. Eble et al. sought to elucidate the mechanism of the Maillardreaction and accordingly conducted controlled initial glycosylation ofRNAase as a model system, which was then examined under varyingconditions. In one aspect, the glycosylated protein material wasisolated and placed in a glucose-free environment and thereby observedto determine the extent of cross-linking.

Eble et al. thereby observed that cross-linking continued to occur notonly with the glycosylated protein but with non-glycosylated proteins aswell. One of the observations noted by Eble et al. was that the reactionbetween glycosylated protein and the protein material appeared to occurat the location on the protein chain of the amino acid lysine.Confirmatory experimentation conducted by Eble et al. in this connectiondemonstrated that free lysine would compete with the lysine on RNAasefor the binding of glycosylated protein. Thus, it might be inferred fromthese data that lysine may serve as an inhibitor of advancedglycosylation; however, this conclusion and the underlying observationsleading to it should be taken in the relatively limited context of themodel system prepared and examined by Eble et al. Clearly, Eble et al.does not appreciate, nor is there a suggestion therein, of thediscoveries that underlie the present invention, with respect to theinhibition of advanced glycosylation of proteins both in vitro and invivo.

The experiments of Eble et al. do not suggest the reactive cleavageproduct mechanism or any other mechanism in the in vivo formation ofadvanced glycosylation endproducts in which glucose is always present.In fact, other investigators support this mechanism to explain theformation of advanced glycosylated endproducts in vivo (see for example;Oimomi et al., Agric. Biol. Chem., 53(6):1727-1728 (1989); and DiabetesResearch and Clinical Practice, 6:311-313 (1989). Accordingly, the useof lysine as an inhibitor in the Eble et al. model system has no bearingupon the utility of the compounds of the present invention in theinhibition of advanced glycosylated endproducts formation in thepresence of glucose in vivo, and the amelioration of complications ofdiabetes and aging.

The compositions useful in the present invention comprise or containagents capable of reacting with the active carbonyl intermediate of anearly glycosylation product. Suitable agents are the compounds ofFormula I of the present invention.

The present invention likewise relates to methods for inhibiting theformation of advanced glycosylation endproducts, which comprisecontacting the target proteins with a composition of the presentinvention. In the instance where the target proteins are contained infoodstuffs, whether of plant or animal origin, these foodstuffs couldhave applied to them by various conventional means a compositioncontaining the present agents.

In the food industry, sulfites were found years ago to inhibit theMaillard reaction and are commonly used in processed and stored foods.Recently, however, sulfites in food have been implicated in severe andeven fatal reactions in asthmatics. As a consequence, the sulfitetreatment of fresh fruits and vegetables has been banned. The mechanismfor the allergic reaction is not known. Accordingly, the presentcompositions and agents offer a nontoxic alternative to sulfites in thetreatment of foods in this manner.

As is apparent from a discussion of the environment of the presentinvention, the present methods and compositions hold the promise forarresting the aging of key proteins both in animals and plants, andconcomitantly, conferring both economic and medical benefits as a resultthereof. In the instance of foodstuffs, the administration of thepresent composition holds the promise for retarding food spoilagethereby making foodstuffs of increased shelf life and greateravailability to consumers. Replacement of currently-used preservatives,such as sulfur dioxide known to cause allergies and asthma in humans,with non-toxic, biocompatible compounds is a further advantage of thepresent invention.

The therapeutic implications of the present invention relate to thearrest of the aging process which has, as indicated earlier, beenidentified in the aging of key proteins by advanced glycosylation andcross-linking. Thus, body proteins, and particularly structural bodyproteins, such as collagen, elastin, lens proteins, nerve proteins,kidney glomerular basement membranes and other extravascular matrixcomponents would all benefit in their longevity and operation from thepractice of the present invention. The present invention thus reducesthe incidence of pathologies involving the entrapment of proteins bycross-linked target proteins, such as retinopathy, cataracts, diabetickidney disease, glomerulosclerosis, peripheral vascular disease,arteriosclerosis obliterans, peripheral neuropathy, stroke,hypertension, atherosclerosis, osteoarthritis, periarticular rigidity,loss of elasticity and wrinkling of skin, stiffening of joints,glomerulonephritis, etc. Likewise, all of these conditions are inevidence in patients afflicted with diabetes mellitus. Thus, the presenttherapeutic method is relevant to treatment of the noted conditions inpatients either of advanced age or those suffering from one of thementioned pathologies.

Protein cross-linking through advanced glycosylation product formationcan decrease solubility of structural proteins such as collagen invessel walls (see Brownlee et al., Science, 232. pp. 1629-1632, (1986)),and can also trap serum proteins, such as lipoproteins to the collagen.Also, this may result in increased permeability of the endothelium andconsequently covalent trapping of extravasated plasma proteins insubendothelial matrix, and reduction in susceptibility of both plasmaand matrix proteins to physiologic degradation by enzymes. (See Brownleeet al., Diabetes, 35, Suppl. 1, p. 42A (1986)). For these reasons, theprogressive occlusion of diabetic vessels induced by chronichyperglycemia has been hypothesized to result from excessive formationof glucose-derived cross-links. Such diabetic macrovascular changes andmicrovascular occlusion can be effectively prevented by chemicalinhibition of advanced glycosylation product formation utilizing acomposition and the methods of the present invention.

Studies indicate that the development of chronic diabetic damage intarget organs is primarily linked to hyperglycemia so that tightmetabolic control would delay or even prevent end-organ damage. SeeNicholls et al., Lab. Invest., 60, No. 4, p. 486 (1989), which discussesthe effects of islet isografting and aminoguanidine in murine diabeticnephropathy. These studies further evidence that aminoguanidinediminishes aortic wall protein cross-linking in diabetic rats andconfirm earlier studies by Brownlee et al., Science, 232, pp. 1629-1632(1986) to this additional target organ of complication of diabetes.Also, an additional study showed the reduction of immunoglobulintrapping in the kidney by aminoguanidine (Brownlee et al., Diabetes, 35,Suppl. 1, p. 42A (1986)).

Further evidence in the streptozotocin-diabetic rat model thataminoguanidine administration intervenes in the development of diabeticnephropathy was presented by Brownlee et al., 1988, supra, with regardto morphologic changes in the kidney which are hallmarks of diabeticrenal disease. These investigators reported that the increasedglomerular basement membrane thickness, a major structural abnormalitycharacteristic of diabetic renal disease, was prevented withaminoguanidine.

Taken together, these data strongly suggest that inhibition of theformation of advanced glycosylation endproducts (AGEs), by the teachingof the present invention, may prevent late, as well as early, structurallesions due to diabetes, as well as changes during aging caused by theformation of AGE's.

Diabetes-induced changes in the deformability of red blood cells,leading to more rigid cell membranes, is another manifestation ofcross-linking and aminoguanidine has been shown to prevent it in vivo.In such studies, New Zealand White rabbits, with induced, long-termdiabetes are used to study the effects of a test compound on red bloodcell (RBC) deformability (df). The test compound is administered at arate of 100 mg/kg by oral gavage to diabetic rabbits (Brown et al.,Presentation of Abstract for Association for Academic MinorityPhysicians, Annual Scientific Meeting (1989)).

Increased cross-linking of collagen in diabetic rats has shown to beprevented by aminoguanidine. Oxlund and Andreassen, "The increase inbiochemical and biomechanical stability of collagen in diabetic rats isprevented by aminoguanidine treatment", European Association for theStudy of Diabetes, Twenty-fifth Annual Meeting, p. 525A, Abstract No.371, 1989 showed the effect when thermal stability of tendon fibers wasassessed by breaking time in a urea bath, as well as mechanicalstrength. Soulis et al., "Aminoguanidine reduces tissue fluorescence butnot albuminuria in diabetic rats". NIH Conference on the MaillardReaction in Aging, Diabetes, and Nutrition, Bethesda, Maryland, Sep.22-23, 1988, page 30) showed the same effect on collagen in the aorta,measured by fluorescence and solubility.

Giambione and Brownlee, "Aminoguanidine Treatment Normalizes IncreasedSteady-state Levels of Laminin B1 mRNA in Kidneys of Long-termStreptozotocin-diabetic Rats" Diabetes, 38, Supplement 2:83A Forty-ninthAnnual Meeting, American Diabetes Association (1989) showed thataminoguanidine treatment to diabetic rats prevents the diabetes-inducedincrease in laminin B₁ mRNA in the kidney. This indicates thataminoguanidine may prevent overproduction of matrix, which leads tobasement membrane thickening and morphologic and functionaldeterioration of vasculature in kidneys and other organs. A furtherconsequence of diabetes is the hyperglycemia-induced matrix bonedifferentiation resulting in decreased bone formation usually associatedwith chronic diabetes. In animal models, diabetes reduces matrix-inducedbone differentiation by 70% (Am. J. Phys., 238 (1980)).

In the instance where the compositions of the present invention areutilized for in vivo or therapeutic purposes, it may be noted that thecompounds or agents used therein are biocompatible. Pharmaceuticalcompositions may be prepared with a therapeutically effective quantityof the agents or compounds of the present invention and may include apharmaceutically acceptable carrier, selected from known materialsutilized for this purpose. Such compositions may be prepared in avariety of forms, depending on the method of administration. Also,various pharmaceutically acceptable addition salts of the compounds ofFormula I may be utilized.

A liquid form would be utilized in the instance where administration isby intravenous, intramuscular or intraperitoneal injection. Whenappropriate, solid dosage forms such as tablets, capsules, or liquiddosage formulations such as solutions and suspensions, etc., may beprepared for oral administration. For topical or dermal application tothe skin or eye, a solution, a lotion or ointment may be formulated withthe agent in a suitable vehicle such as water, ethanol, propyleneglycol, perhaps including a carrier to aid in penetration into the skinor eye. For example, a topical preparation could include up to about 10%of the compound of Formula I. Other suitable forms for administration toother body tissues are also contemplated.

In the instance where the present method has therapeutic application,the animal host intended for treatment may have administered to it aquantity of one or more of the agents, in a suitable pharmaceuticalform. Administration may be accomplished by known techniques, such asoral, topical and parenteral techniques such as intradermal,subcutaneous, intravenous or intraperitoneal injection, as well as byother conventional means. Administration of the agents may take placeover an extended period of time at a dosage level of, for example, up toabout 25 mg/kg.

As noted earlier, the invention also extends to a method of inhibitingthe discoloration of teeth resulting from nonenzymatic browning in theoral cavity which comprises administration to a subject in need of suchtherapy an amount effective to inhibit the formation of advancedglycosylation endproducts of a composition comprising an agent of thestructural Formula I.

The nonenzymatic browning reaction which occurs in the oral cavityresults in the discoloration of teeth. Presently used anti-plaque agentsaccelerate this nonenzymatic browning reaction and further the stainingof the teeth. Recently, a class of cationic anti-microbial agents withremarkable anti-plaque properties have been formulated in oral rinsesfor regular use to kill bacteria in the mouth. These agents, thecationic antiseptics, include such agents as alexidine, cetyl pyridiniumchloride, chlorhexidine gluconate, hexetidine, and benzalkoniumchloride.

Tooth staining by chlorhexidine and other anti-plaque agents apparentlyresults from the enhancement of the Maillard reaction. Nordbo, J. Dent.Res.. 58, p. 1429 (1979) reported that chlorhexidine and benzalkoniumchloride catalyze browning reactions in vitro. Chlorhexidine added tomixtures containing a sugar derivative and a source of amino groupsunderwent increased color formation, attributed to the Maillardreaction. It is also known that use of chlorhexidine results in anincreased dental pellicle. Nordbo proposed that chlorhexidine resultedin tooth staining in two ways: first, by increasing formation ofpellicle which contains more amino groups, and secondly, by catalysis ofthe Maillard reaction leading to colored products.

In accordance with this method, the compounds of Formula I areformulated into compositions adapted for use in the oral cavity.Particularly suitable formulations are oral rinses and toothpastesincorporating the active agent.

In the practice of this invention, conventional formulating techniquesare utilized with nontoxic, pharmaceutically acceptable carrierstypically utilized in the amounts and combinations that are well-knownfor the formulation of such oral rinses and toothpastes.

The agent of Formula I is formulated in compositions in an amounteffective to inhibit the formation of advanced glycosylationendproducts. This amount will, of course, vary with the particular agentbeing utilized and the particular dosage form, but typically is in therange of 0.01% to 1.0%, by weight, of the particular formulation.

Additionally, since the agents of the aforesaid method are concentratedin the salivary glands upon oral ingestion or parenteral administration,they can be so administered. This concentration in the salivary glandsresults in their secretion into saliva, the net result being that theyare functionally placed in the oral cavity where they can effect theirdesired method. For such administration, the particular agent can beformulated in any conventional oral or parenteral dosage form. Aparticularly desirable dosage form is the incorporation of the agentinto a vitamin tablet or fluoride tablet so as to maximize patient, andparticularly juvenile patient, compliance.

The novel compounds of Formula I of the present invention areconveniently prepared by reacting the appropriate starting material ofthe formula ##STR3## wherein n and R₁ are as hereinbefore defined and Yis an appropriate leaving group with an amino compound of the formula

    NH.sub.2 --R.sub.2                                         (III)

wherein R₂ is as hereinbefore defined, in a protic solvent. Typically,the solvent is an alcohol such as ethanol or isopropanol. The reactionis conducted at temperatures from room temperature to reflux (dependingupon the chosen solvent). Typical reaction times vary from 1-12 hours.Appropriate leaving groups are those such as methylthio groups, chloro,bromo or iodo atoms, and the like.

The starting materials of Formulae II and III are generally known fromthe literature (see, for instance, U.S. Pat. No. 4,908,446). However, inthe case wherein R₁ is a hydroxyethyl group, the intermediate of FormulaII is conveniently prepared by reaction of either(hydroxyethyl)ethylenediamine (for n=1) or 2-(3-aminopropylamino)ethanol (for n=2) with carbon disulfide. The resulting1-(2-hydroxylethyl)-2-imidazolinethione (n=1) or1-(2-hydroxyethyl)-3,4,5,6-tetrahydropyrimidine-2-thione (n=2) is thenreacted to with methyl methanesulfonate to form the necessaryintermediate of Formula II.

The following examples are illustrative of the compounds, compositionsand methods of the present invention.

EXAMPLE I

2-(3-Aminopropylamino)ethanol (118 ml) is dissolved in 1:1 ethylalcohol-water (340 ml) and heated to 40°-50° C. Carbon disulfide (60 ml)is added dropwise over a period of about 15 minutes. An exothermicreaction occurs after about half of the amount of carbon disulfide isadded. After the addition of all the carbon disulfide, the mixture isrefluxed for about 1 hour. Concentrated hydrochloric acid (8 ml) is thenadded, and the reaction mixture is refluxed overnight. After cooling inan ice bath, the crystalline solid which separates is filtered, washedwith isopropyl alcohol and t-butyl methyl ether, and dried to give 129-4g of 1-(2-hydroxyethyl)-3,4,5,6-tetrahydropyrimidine-2-thione, meltingpoint 112°-114° C.

Then, 1-(2-hydroxyethyl)-3,4,5,6-tetrahydropyrimidine-2-thione (121.2 g)and methyl methanesulfonate (91.51 g) are heated at reflux in ethylalcohol (200 ml) for about 2 hours and then allowed to cool to roomtemperature. The mixture was diluted with t-butyl methyl ether (2liters). An oil separates which crystallizes on keeping in freezer at-20° C. Filtration and drying yields 145.7 g of1-(2-hydroxyethyl)-2-methylthio-3,4,5,6-tetrahydropyrimidine mesylate,melting point 61.7°-62.2° C.

1-(2-hydroxyethyl)-2-methylthio-3,4,5,6-tetrahydropyrimidine mesylate(87.2 g) is suspended in ethyl alcohol (150 ml) and treated withanhydrous hydrazine (22 mL). The mixture becomes clear within a fewminutes. It is stirred at room temperature overnight and then dilutedwith t-butyl methyl ether (2 liters). The solution is kept in thefreezer at -20° C. until the solid separates, filters and dries.Recrystallization from isopropyl alcohol and t-butyl methyl ether gives68.12 g of 1-(2-hydroxyethyl)- 2-hydrazino-3,4,5,6-tetrahydropyrimidinemesylate, melting point 82.2°-84.4° C.

EXAMPLE 2

(Hydroxyethyl)ethylenediamine (25 ml) is dissolved in 1:1 ethylalcohol-water (85 mL) and heated to 40°-50° C. Carbon disulfide (16.5ml) is added dropwise over a period of about 15 minutes. After abouthalf of the amount is added, the exothermic reaction starts and thereaction mixture solidifies. After the addition of all of the carbondisulfide, the mixture is refluxed gently for about 1 hour. Concentratedhydrochloric acid (2 ml) is then added and the reaction mixture isrefluxed overnight. The solid which separates after the addition ofcarbon disulfide redissolves. The solution is cooled in an ice bath, andthe solid separates, is filtered out, washed well with isopropyl alcoholand t-butyl methyl ether and dried to give 23.3 g of1-(2-hydroxyethyl)-2-imidazolinethione, melting point 137°-138° C.

Twenty grams of 1-(2-hydroxyethyl)-2-imidazolinethione and methylmethanesulfonate (12.75 ml) are dissolved in ethyl alcohol (90 mL) andrefluxed for 2 hours. The mixture is cooled to room temperature anddiluted with t-butyl methyl ether (1 liter). An oil separates whichsolidifies on keeping the mixture in the freezer at -20° C. for a fewdays. The solid is then filtered, washed with butyl methyl ether anddried to give 1-(2-hydroxyethyl)-2-methylthioimidazoline mesylate,melting point 78.9°-81.5° C.

1-(2-Hydroxyethyl)-2-methylthioimidazoline mesylate (35 g, 0.136 mole)is suspended in ethyl alcohol (80 ml) and treated with anhydroushydrazine (9.3 ml, 0.296 mole). The mixture became clear immediately.The solution is stirred at room temperature overnight and then dilutedwith t-butyl methyl ether (1 liter). On storage of the mixture in thefreezer at -20° C., crystals separated, which are filtered, washed wellwith t-butyl methyl ether and dried. Recrystallization from isopropylalcohol and t-butyl methyl ether gives1-(2-hydroxyethyl)-2-hydrazinoimidazoline mesylate, melting point85°-88° C.

EXAMPLE 3

1-Amino-2-methylthio-2-imidazoline (U.S. Pat. No. 4,908,446) (3.03 g)and isopropanol (7.5 ml) were placed in a 25 ml round bottom flask, and2-hydroxyethylhydrazine (1.52 g) was added dropwise. The mixture isheated to about 40° C. to give a solution. The heat is removed andstirring is continued at 25° C. overnight. The deposited crystals arefiltered out, washed with isopropanol and air dried. Recrystallizationfrom isopropanol gives 0.74 g of1-amino-2-[2-(2-hydroxyethyl)-hydrazino]2-imidazoline tosylate ascolorless crystals, melting point 26.9°-127.2°.

EXAMPLE 4

1-Amino-2-methylthio-2-imidazoline tosylate (3.03 g) and isopropanol(7.5 ml) are placed in a 25 ml round bottom flask, then 2-aminoethanol(1.22 g) is added. The mixture is heated to about 40° C. to give asolution. After stirring overnight at 25° C., the mixture isconcentrated to a thick colorless oil. Crystals were form upon storageat -20° C. Trituration with 10 ml isopropanol and filtration gives 2.68g (84.8%) of 1-amino-2-(2-hydroxyethyl)-2-imidazoline tosylate ascolorless crystals with mp 91.8°-92.7° C.

EXAMPLE 5

1-Amino-2-methylthio-2-imidazoline tosylate (1 g) and4-(2-aminoethyl)morpholine (0.47 g) are dissolved in ethyl alcohol (10ml) and refluxed overnight. The reaction mixture is cooled to roomtemperature and diluted with t-butyl methyl ether (75 ml). The whitesolid which separates, is filtered, washed well with t-butyl methylether, and dried. Crystallization from isopropyl alcohol gives 940 mg ofcolorless crystals of1-amino-2-(4-[2-aminoethyl]morpholino)-2-imidazoline tosylate, meltingpoint 139.9°-141° C.

Following analogous procedures, the following aminoalkyl imidazolinederivatives are prepared (substituting for 4-(2-aminoethyl)morpholinethe following reagents:

From 4-(3-aminopropyl)morpholine, the compound1-amino-2-(4-[3-aminopropyl]morpholino)-2-imidazoline tosylate, meltingpoint 154°-155° C.

From 1-(3-aminopropyl)-4-methylpiperazine, the compound1-amino-2-(1-[3-aminopropyl]-4-methylpiperazino)-2-imidazoline tosylate,melting point 127°-129° C.

From 3-dimethylaminopropylamine, the compound1-amino-2-[3-(dimethylamino)propylamino]-2-imidazoline tosylate, meltingpoint 112°-114° C.

From 3-ethoxypropylamine, the compound1-amino-2-(3-ethoxypropylamino)-2-imidazoline tosylate, melting point106°-107° C.

From 1-(3-aminopropyl)imidazole, the compound1-amino-2-[3-(1-imidazolyl)propyl]amino-2-imidazoline tosylate, meltingpoint 123°-125° C.

From 2-methoxyethylamine, the compound1-amino-2-(2-methoxyethyl)amino-2-imidazoline tosylate, melting point94.2°-97.2°.

From 3-isopropoxypropylamine, the compound1-amino-2-(3-methylthiopropylamino)-2-imidazoline tosylate, meltingpoint 98°-100° C.

From 1-amino-3-(N-piperidino)propane, the compound1-amino-2-[1-amino-3-(N-piperidino)propyl]-2-imidazoline tosylate,melting point 137°-139.7°.

From N,N,2,2-tetramethyl-1,3-propanediamine, the compound1-amino-2-(N,N,2,2-tetramethyl-1,3-propanediamino)-2-imidazolinetosylate, melting point 101.9°-102.9°.

From neopentylamine, the compound1-amino-2-(neopentylamino)-2-imidazoline tosylate, melting point166.4°-167.5° C.

EXAMPLE 6

The following method was used to evaluate the ability of the compoundsof the present invention to inhibit glucose-mediated development offluorescence of bovine serum albumin (BSA), a measure of cross-linking.Compounds were incubated under aseptic conditions at a concentration of1 mM with 400 mM glucose and 100 rg/mL BSA in a 1.5 M sodium phosphatebuffer, pH 7.4.

Samples of the incubation mixture were taken immediately and after 1week incubation at 37 C for measurement of fluorescence. For each testcompound, control incubations in buffer were made of compound alone (C),compound plus glucose (G+C), and compound plus BSA (B+C). An additionalset of incubations of glucose and BSA (B+G) were prepared as thebaseline controls against which were measured the ability of thecompounds to inhibit. Each incubation was made in triplicate.

Fluorescence (excitation, 370 nm; emission, 440 nm) was measured on eachsample after a 100-fold dilution in distilled water.

The % inhibition of browning of each test compound was calculated asfollows. Each F represents the fluorescence measurement of that sampleafter 1 week incubation less its fluorescence before incubation.##EQU1## where B=BSA, G=glucose, and C-test compound.

Percent inhibition of browning by various test compounds at 1 mM:

    ______________________________________                                        0%            no inhibitor                                                    68.1%         1-amino-2-[2-(2-hydroxyethyl)hydrazino]-2-                                    imidazoline tosylate;                                           61%           1-amino-2-(2-hydroxyethylamino)-2-                                            imidazoline tosylate;                                           38.5%         1-(2-hydroxyethyl)-2-hydrazino-1,4,5,6-                                       tetrahydropyrimidine                                                          methanesulfonate;                                               45.1%         1-(2-hydroxyethyl)-2-hydrazino-2-                                             imidazoline methanesulfonate;                                   5.9%          1-amino-2-([3-(4-morpholino)propyl]                                           amino)imidazoline tosylate;                                     16.7%         1-amino-2-([3-(dimethylamino)propyl]                                          amino)imidazoline tosylate;                                     19.9%         1-amino-2-([3-(1-imidazolyl)propyl]                                           amino)imidazoline tosylate;                                     ______________________________________                                    

The above experiments suggest that this type of drug therapy may havebenefit in reducing the pathology associated with the advancedglycosylation of proteins and the formation of cross-links betweenproteins and other macromolecules. Drug therapy may be used to preventthe increased trapping and cross-linking of proteins that occurs indiabetes and aging which leads to sequelae such as retinal damage, andextra-vascularly, damage to tendons, ligaments and other joints. Thistherapy might retard atherosclerosis and connective tissue changes thatoccur with diabetes and aging. Both topical, oral, and parenteral routesof administration to provide therapy locally and systemically arecontemplated.

EXAMPLE 7

    ______________________________________                                        Tablet            mg/tablet                                                   ______________________________________                                        Compound of Formula I                                                                           50                                                          Starch            50                                                          Mannitol          75                                                          Magnesium stearate                                                                               2                                                          Stearic acid       5                                                          ______________________________________                                    

The compound, a portion of the starch and the lactose are combined andwet granulated with starch paste. The wet granulation is placed on traysand allowed to dry overnight at a temperature of 45° C. The driedgranulation is comminuted in a comminutor to a particle size ofapproximately 20 mesh. Magnesium stearate, stearic acid and the balanceof the starch are added and the entire mix blended prior to compressionon a suitable tablet press. The tablets are compressed at a weight of232 mg. using a 11/32" punch with a hardness of 4 kg. These tablets willdisintegrate within a half hour according to the method described in USPXVI.

EXAMPLE 8

    ______________________________________                                        Lotion                  mg/g                                                  ______________________________________                                        Compound of Formula I    1.0                                                  Ethyl alcohol           400.0                                                 Polyethylene glycol 400 300.0                                                 Hydroxypropyl cellulose  5.0                                                  Propylene glycol to make                                                                               1.0 g                                                ______________________________________                                    

EXAMPLE 8

    ______________________________________                                        Oral Rinse                                                                    ______________________________________                                        Compound of Formula I:  1.4%                                                  Chlorhexidine gluconate 0.12%                                                 Ethanol                 11.6%                                                 Sodium saccharin        0.15%                                                 FD&C Blue No. 1         0.001%                                                Peppermint Oil          0.5%                                                  Glycerine               10.0%                                                 Tween 60                0.3%                                                  Water to                100%                                                  ______________________________________                                    

EXAMPLE 9

    ______________________________________                                        Toothpaste                                                                    ______________________________________                                        Compound of Formula I:    5.5%                                                Sorbitol, 70% in water    25%                                                 Sodium saccharin          0.15%                                               Sodium lauryl sulfate     1.75%                                               Carbopol 934, 6% dispersion in water                                                                    15%                                                 Oil of Spearmint          1.0%                                                Sodium hydroxide, 50% in water                                                                          0.76%                                               Dibasic calcium phosphate dihydrate                                                                     45%                                                 Water to                  100%                                                ______________________________________                                    

EXAMPLE 11

To further study the ability of inhibitors of nonenzymatic browning toprevent the discoloration of protein on a surface, such as that whichoccurs on the tooth surface, the following surface browning experimentis performed. As a substitute for a pellicle-covered tooth surface,unexposed and developed photographic paper is used to provide a fixedprotein (gelatin, i.e., collagen) surface on a paper backing. Fivemillimeter circles are punched and immersed for one week at 50° C. in asolution of 100 mM glucose-6-phosphate in a 0.5 M phosphate buffer, pH7.4, containing 3 mM sodium azide. Glucose-6-phosphate is a sugarcapable of participating in nonenzymatic browning at a more rapid ratethan glucose. In addition to the glucose-6-phosphate, chlorhexidineand/or a compound of Formula I are included. After incubation, thegelatin/paper disks are rinsed with water, observed for brown color, andphotographed.

Incubation of the disks in glucose-6-phosphate alone shows slight browncolor versus disks soaked in buffer alone. Inclusion of chlorhexidine(in the form of Peridex® at a final concentration of 0.04%chlorhexidine) shows significant browning. Addition of a compound ofFormula I to the chlorhexidine inhibits browning of the gelatin, as doesinclusion of a compound of Formula I in the absence of chlorhexidine.

The slight brown color formed by the action of glucose-6-phosphate onthe gelatin surface alone and its prevention by a compound of Formula Idemonstrates the utility of the present invention in preventingnonenzymatic browning of tooth surfaces. The enhanced browning in thepresence of chlorhexidine and its prevention with a compound of FormulaI demonstrates the utility of the present invention in preventing theanti-plaque agent-enhanced nonenzymatic browning which occurs withchlorhexidine.

This invention may be embodied in other forms or carried out in otherways without departing from the spirit or essential characteristicsthereof. The present disclosure is therefore to be considered as in allrespects illustrative and not restrictive, the scope of the inventionbeing indicated by the appended Claims, and all changes which comewithin the meaning and range of equivalency are intended to be embracedtherein.

What is claimed is:
 1. A compound of the formula ##STR4## wherein n is1;R₁ is a hydroxyethyl or an amino group; R₂ is an amino, a hydroxyloweralkylamino, a lower alkyl group, or a group of the formula

    alk-Y

wherein alk is a lower alkylene group and Y is selected from the groupconsisting of hydroxy, lower alkoxy, lower alkylthio and heterocyclicgroups selected from the group consisting of imidazolyl, morpholino,piperidino, piperazinyl, methylpiperazinyl and hexamethylenimino withthe proviso that when R₁ is a hydroxy ethyl group, then R₂ is an aminogroup; and the biocompatible and pharmaceutically acceptable saltsthereof.
 2. A compound according to claim 1 wherein R₁ is a hydroxyethylgroup.
 3. A compound according to claim 1 which is1-(2-hydroxyethyl)-2-hydrazino-2-imidazoline methanesulfonate or anotherpharmaceutically acceptable salt thereof.
 4. A compound according toclaim 1 wherein R₁ is an amino group.
 5. A compound according to claim 4wherein R₂ is a hydroxylower alkyl group.
 6. A compound according toclaim 5 which is 1-amino-2-[2-(2-hydroxyethyl)hydrazino]-2-imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 7. Acompound according to claim 4 wherein R₂ is a lower alkyl group.
 8. Acompound according to claim 7 which is1-amino-2-(neopentylamino)-2-imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 9. A compound according toclaim 4 wherein R₂ is a group of the formula

    alk-Y

wherein alk is a lower alkylene group and Y is selected from the groupconsisting of hydroxy, lower alkoxy, lower alkyl thio, lower alkylaminoand heterocyclic groups selected from the group consisting ofimidazolyl, morpholino, piperidino, piperazinyl, methylpiperazinyl andhexamethylenimino.
 10. A compound according to claim 9 which is1-amino-2-(2-hydroxyethylamino)-2-imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 11. A compound according toclaim 9 which is 1-amino-2-[(3-ethoxypropyl)amino]imidazoline tosylateor another pharmaceutically acceptable salt thereof.
 12. A compoundaccording to claim 9 which is1-amino-2-(2-methoxyethylamino)-2-imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 13. A compound according toclaim 9 which is 1-amino-2-(3-isopropoxypropyl amino)-2-imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 14. Acompound according to claim 9 which is1-amino-2-(3-methylthiopropylamino)-2-imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 15. A compound according toclaim 9 which is 1-amino-2-([3-(dimethylamino)propyl]amino)imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 16. Acompound according to claim 9 which is1-amino-2-[2,2-dimethyl-3-(dimethylamino)propylamino]-2-imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 17. Acompound according to claim 9 which is1-amino-2-([2-(4-morpholino)ethyl]amino)imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 18. A compound according toclaim 9 which is 1-amino-2-(2-([3-(4-morpholino)propyl]amino)imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 19. Acompound according to claim 9 which is1-amino-2-([3-(4-methylpiperazin-1-yl)propyl]amino)imidazoline tosylateor another pharmaceutically acceptable salt thereof.
 20. A compoundaccording to claim 9 which is1-amino-2-([3-(1-imidazolyl)propyl]amino)imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 21. A compound according toclaim 9 which is 1-amino-2-[3-(1-piperidino)propylamino)imidazolinetoluenesulfonate or another pharmaceutically acceptable salt thereof.22. A composition for inhibiting the advanced glycosylation of a targetprotein comprising an effective amount of a compound selected from thegroup consisting of compounds of the formula ##STR5## wherein n is 1;R₁is a hydroxyethyl or an amino group; R₂ is an amino, a hydroxyloweralkylamino, a lower alkyl group, or a group of the formula

    alk-Y

wherein alk is a lower alkylene group and Y is selected from the groupconsisting of hydroxy, lower alkoxy, lower alkylthio and heterocyclicgroups selected from the group consisting of imidazolyl, morpholino,piperidino, piperazinyl, methylpiperazinyl and hexamethylenimino withthe proviso that when R₁ is a hydroxy ethyl group, then R₂ is an aminogroup; and the biocompatible and pharmaceutically acceptable saltsthereof.
 23. A pharmaceutical composition for administration to ananimal to inhibit the advanced glycosylation of a target protein withinsaid animal, comprising a pharmaceutically effective amount of acompound selected from the group consisting of compounds of the formula##STR6## wherein n is 1;R₁ is a hydroxyethyl or an amino group; R₂ is anamino, a hydroxylower alkylamino, a lower alkyl group, or a group of theformula

    alk-Y

wherein alk is a lower alkylene group and Y is selected from the groupconsisting of hydroxy, lower alkoxy, lower alkylthio and heterocyclicgroups selected from the group consisting of imidazolyl, morpholino,piperidino, piperazinyl, methylpiperazinyl and hexamethylenimino withthe proviso that when R is a hydroxy ethyl group, then R₂ is an aminogroup; and the biocompatible and pharmaceutically acceptable saltsthereof.
 24. The composition of claim 23 wherein said compound has theformula wherein R₁ is an hydroxyethyl group.
 25. The composition ofclaim 24 wherein said compound is1-(2-hydroxyethyl)-2-hydrazino-2-imidazoline methanesulfonate or anotherpharmaceutically acceptable salt thereof.
 26. The composition of claim23 wherein said compound has the formula wherein R₁ is an amino group.27. The composition of claim 26 wherein said compound is1-amino-2-[2-(2-hydroxyethyl)hydrazino]-2-imidazolinetosylate or anotherpharmaceutically acceptable salt thereof.
 28. The composition of claim26 wherein said compound is 1-amino-2-(neopentylamino)-2-imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 29. Thecomposition of claim 26 wherein said compound is1-amino-2-(2-hydroxyethylamino)-2-imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 30. The composition of claim26 wherein said compound is 1-amino-2-[(3-ethoxypropyl)amino]imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 31. Thecomposition of claim 26 wherein said compound is1-amino-2-(2-methoxyethylamino)-2-imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 32. The composition of claim26 wherein said compound is1-amino-2-(3-isopropoxypropylamino)-2-imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 33. The composition of claim26 wherein said compound is1-amino-2-(3-methylthiopropylamino)-2-imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 34. The composition of claim26 wherein said compound is1-amino-2-([3-(dimethylamino)propyl]amino)imidazoline tosylate oranother pharmaceutically acceptable salt thereof.
 35. The composition ofclaim 26 wherein said compound is1-amino-2-[2,2-dimethyl-3-(dimethylamino)propylamino]-2-imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 36. Thecomposition of claim 26 wherein said compound is1-amino-2-([2-(4-morpholino)ethyl]amino)imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 37. The composition of claim26 wherein said compound is1-amino-2-([3-(4-morpholino)propyl]amino)imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 38. The composition of claim26 wherein said compound is1-amino-2-([3-(4-methylpiperazin-1-yl)propyl]amino)imidazoline tosylateor another pharmaceutically acceptable salt thereof.
 39. The compositionof claim 26 wherein said compound is1-amino-2-([3-(1-imidazolyl)propyl]amino)imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 40. The composition of claim26 wherein said compound is1-amino-2-([3-(1-piperidino)propyl]amino)imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 41. A method for inhibitingthe advanced glycosylation of a target protein comprising contacting thetarget protein with an effective amount of composition comprising acompound selected from the group consisting of compounds of the formula##STR7## wherein n is 1;R₁ is a hydroxyethyl or an amino group; R₂ is anamino, a hydroxylower alkylamino, a lower alkyl group, or a group of theformula

    alk-Y

wherein alk is a lower alkylene group and Y is selected from the groupconsisting of hydroxy, lower alkoxy, lower alkylthio and heterocyclicgroups selected from the group consisting of imidazolyl, morpholino,piperidino, piperazinyl, methylpiperazinyl and hexamethylenimino withthe proviso that when R₁ is a hydroxy ethyl group, then R₂ is an aminogroup; and the biocompatible and pharmaceutically acceptable saltsthereof.
 42. A method for treating an animal to inhibit the formation ofadvanced glycosylation endproducts of a target protein within saidanimal, said method comprising administering an effective amount of apharmaceutical composition, said pharmaceutical composition comprising acompound selected from the group consisting of compounds of the formula##STR8## wherein n is 1;R₁ is a hydroxyethyl or an amino group; R₂ is anamino, a hydroxylower alkylamino, a lower alkyl group, or a group of theformula

    alk-Y

wherein alk is a lower alkylene group and Y is selected from the groupconsisting of hydroxy, lower alkoxy, lower alkylthio and heterocyclicgroups selected from the group consisting of imidazolyl, morpholino,piperidino, piperazinyl, methylpiperazinyl and hexamethylenimino withthe proviso that when R is a hydroxy ethyl group, then R₂ is an aminogroup; and the biocompatible and pharmaceutically acceptable saltsthereof.
 43. The method of claim 42 wherein said compound has theformula wherein R₁ is a hydroxyethyl group.
 44. The method of claim 43wherein said compound is 1-(2-hydroxyethyl)-2-hydrazino-2-imidazolinemethanesulfonate or another pharmaceutically acceptable salt thereof.45. The method of claim 42 wherein said compound has the formula whereinR₁ is an amino group.
 46. The method of claim 45 wherein said compoundis 1-amino-2-[2-(2-hydroxyethyl)hydrazino]-2-imidazoline tosylate oranother pharmaceutically acceptable salt thereof.
 47. The method ofclaim 45 wherein said compound is1-amino-2-(neopentylamino)-2-imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 48. The method of claim 45wherein said compound is 1-amino-2-(2-hydroxyethylamino)-2-imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 49. Themethod of claim 45 wherein said compound is1-amino-2-[(3-ethoxypropyl)amino]imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 50. The method of claim 45wherein said compound is 1-amino-2-(2-methoxyethylamino)-2-imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 51. Themethod of claim 45 wherein said compound is1-amino-2-(3-isopropoxypropylamino)-2-imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 52. The method of claim 45wherein said compound is1-amino-2-(3-methylthiopropylamino)-2-imidazoline tosyltae or anotherpharmaceutically acceptable salt thereof.
 53. The method of claim 45wherein said compound is1-amino-2-([3-(dimethylamino)propyl]amino)imidazoline tosylate oranother pharmaceutically acceptable salt thereof.
 54. The method ofclaim 45 wherein said compound is1-amino-2-[2,2-dimethyl-3-(dimethylamino)propylamino]-2-imidazolinetosylate or another pharmaceutically acceptable salt thereof.
 55. Themethod of claim 45 wherein said compound is1-amino-2-([2-(4-morpholino)ethyl]amino)imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 56. The method of claim 45wherein said compound is1-amino-2-([3-(4-morpholino)propyl]amino)imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 57. The method of claim 47wherein said compound is1-amino-2-([3-(4-methylpiperazin-1-yl)propyl]amino)imidazoline tosylateor another pharmaceutically acceptable salt thereof.
 58. The method ofclaim 45 wherein said compound is1-amino-2-([3-(1-imidazolyl)propyl]amino)imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 59. The method of claim 45wherein said compound is1-amino-2-([3-(1-piperidino)propyl]amino)imidazoline tosylate or anotherpharmaceutically acceptable salt thereof.
 60. A method of inhibiting thediscoloration of teeth resulting from non-enzymatic browning in the oralcavity which comprises administration of an amount effective to inhibitthe formation of advanced glycosylation endproducts of a compositioncomprising a compound selected from the group consisting of compounds ofthe formula ##STR9## wherein n is 1;R₁ is a hydroxyethyl or an aminogroup; R₂ is an amino, a hydroxylower alkylamino, a lower alkyl group,or a group of the formula

    alk-Y

wherein alk is a lower alkylene group and Y is selected from the groupconsisting of hydroxy, lower alkoxy, lower alkylthio and heterocyclicgroups selected from the group consisting of imidazolyl, morpholino,piperidino, piperazinyl, methylpiperazinyl and hexamethylenimino withthe proviso that when R₁ is a hydroxy ethyl group, then R₂ is an aminogroup; and the biocompatible and pharmaceutically acceptable saltsthereof.